Investigations of ischemic injury and cardiac protection over the past two decades have greatly advanced our understanding of the cellular events that mediate these biological processes. However, significant controversies exist with regard to the specific molecules and signaling mechanisms that underlie cardiac cell death and survival. We believe that the use of divergent models in isolation and the analysis of individual proteins-in the absence of thorough investigation of the associated molecules and their respective interactions at the subproteomic level-has hampered accurate characterization of how proteins contribute to a phenotype in vivo. These issues have in turn negatively impacted the translation of basic science advances of the past 20 years to improved treatment of ischemic heart disease. This Program Project takes a novel approach to address the aforementioned limitations and therefore affords great opportunities for conceptual and mechanistic advancement of the field of myocardial ischemia and cardiac protection. We feel that a significant paradigm shift is required to accelerate translational research in the area of myocardial ischemia. Accordingly, instead of relying on one particular experimental model, our unique team of investigators enables the characterization of the role of specific proteins using an array of models at the system, organ, cell and organelle levels. Instead of focusing the study on one pathway and/or a single molecule in isolation, the novel proteomic technology platform provided by the Proteomic Core enables systematic examination of a protein and its associating partners, i.e., the subproteome in which the protein functions in the setting of ischemic injury and cardiac protection. Moreover, subsequent to subproteome mapping studies, our investigator team will engage in a rigorous target validation process. This validation process will be facilitated by the state-of-the-art techniques assembled in individual Projects as well as the Heart Biology Core. These techniques include cardiac-targeted inducible transgenesis, high resolution confocal microscopy, and integrative physiology, which in combination will allow for comprehensive and unequivocal investigation of the mechanistic nature of ischemic injury and protection. Importantly, these studies will target subproteomes involved in physiologic phenotypes and therefore the findings will facilitate an understanding of the discrete molecular context in which specific molecules elicit their cellular effects. These investigations will elucidate more accurate targets for therapeutic intervention and thus will be inherently more translatable to the pre-clinical arena.